JP2007029096A - Analytical method and apparatus - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide a method for conventionally allowing fast amplification of a nucleic acid. <P>SOLUTION: The nucleic acid amplification method comprises a step to form a liquid mixture containing a solid particle having high first energy source absorption power compared with the nucleic acid and a liquid containing a reagent necessary for the amplification of the nucleic acid, and a step to introduce the first energy in the form of pulses of 0.001-10 sec for a period sufficient for heating the solid particle to ≥90°C. The invention further provides a heating method containing a step to heat the mixture simultaneously and/or continuously with different kind of heat sources. <P>COPYRIGHT: (C)2007,JPO&INPIT

Description

  The subject of the present invention is a method for amplifying nucleic acids, a method for treating a liquid, an apparatus for heating a liquid in a container, and a system for heating a liquid in a container.

  The invention is particularly useful in the medical field and requires a reliable analysis of the sample for the components contained therein. Chemical reactions that require heating are well known, for example, from Molecular Diagnostics, where nucleic acids are denatured by applying heat at the hybrid's melting temperature anomalies, i.e., from double-stranded hybrids. It is known to become a main chain.

  A method that uses a reaction cycle that includes such a denaturation step is the polymerase chain reaction (PCR). The technology has revolutionized the field of nucleic acid processing, particularly the field of nucleic acid analysis, provided by means for increasing the amount of nucleic acid of a particular sequence from a negligible amount to a detectable amount. PCR is described in EP 0 201 184 and EP 0 200 362.

  Methods for heating the composition in question are also known. For example, US 2002/0061588 describes a method of heating a nucleic acid by attaching the nucleic acid to a nanoparticle and applying energy to the nanoparticle. This heating denatures the nucleic acid hybrid on the surface of the modulator and dissociates one of the strands into the surrounding liquid. However, the method is very inefficient with respect to heating and amplification.

  US 2004/0129555 describes a method for heating a mixture containing a dye using a pulsed laser.

  U.S. Pat. No. 6,633,785 describes a method for heating microtubes using either resistance heating or induction heating.

  The above prior art methods do not provide temperature conditions for effective amplification of nucleic acids.

SUMMARY OF THE INVENTION A first object of the present invention is a liquid mixture comprising nucleic acids and solid particles having a higher first energy source absorption capacity than the liquid, the mixture comprising the reagents necessary for amplifying the nucleic acids. Providing a first energy in a pulse sufficient for heating the solid particles to a temperature of 90 ° C. or higher, 0.001 to 10 seconds, preferably 0.001 to 1 second. A method for nucleic acid amplification comprising:

  The second object of the present invention is the step of providing the liquid and solid particles in a container, and the step of heating the mixture, wherein the mixture is simultaneously and / or continuously heated by different types of heating sources. A method for heating a liquid comprising.

  Another subject of the present invention is a system for heating a mixture comprising a mixture and a device comprising one or more chambers for containing a first heating source and a second heating source. Where the heating source is located in the system to be effective for heating the contents of the chamber of the device when placed in a container of the device.

  A further subject of the invention is for heating one or more mixtures comprising a container for receiving a device comprising a chamber for containing said mixture, a first heating source and a second heating source. An instrument, wherein the first and second heating sources are located within the instrument to be effective for heating the contents of the chamber of the apparatus when placed in the receptacle.

Detailed Description of the Invention Methods for the amplification of nucleic acids are known. These are intended to create large quantities of nucleic acids based on the initial presence of the target nucleic acid that serves as a template for the activity of an enzyme capable of replicating the base sequence in the target nucleic acid. The replicon itself is used as a target for the replication of the sequence, preferably the base sequence that was already the subject of the first replication. Therefore, a large amount of nucleic acid having the same sequence is created.

  A particularly well-established method for nucleic acid amplification is the polymerase chain reaction (PCR) method disclosed in EP 200362. In the method, the reaction mixture is subjected to repeated cycles of a temperature profile, which anneals the primer with the target gene, extends the annealed primer using the target nucleic acid as a template, and generates an extension from the template. It is adapted to separate things.

  In the first step, a liquid mixture is provided comprising nucleic acids and solid particles having a higher first energy source absorption capacity than the liquid.

  The liquid may be any liquid containing nucleic acid to be amplified. Furthermore, the liquid contains reagents necessary for the amplification of the nucleic acid. These reagents are well known for each amplification method, and preferably an agent for extending the primer, preferably a template-dependent DNA- or RNA-polymerase, and a building block attached to the primer for extension; For example, it contains nucleotides. In addition, the mixture will contain reagents useful for establishing conditions for the extension reaction, such as buffers and cofactors of the enzymes used, such as salts.

  Importantly, however, the mixture contains solid particles. Such solid particles are inert to amplification conditions. That is, they are not destroyed, especially at the temperatures used in the above method. These particles are further characterized by their ability to absorb energy from a first energy source superior to liquids. Obviously, the composition of the particles depends on the energy source.

  The first component of the system according to the present invention is the first source of energy. Preferably, the energy from the first source is transmitted in a manner that does not involve any direct contact between the energy source and the particles. This type of energy transfer is referred to as non-contact energy transfer below. Non-contact energy transfer is performed by electromagnetic radiation, more preferably electromagnetic radiation in the frequency range of 1 kHz to about 50 GHz, most preferably radio frequency (RF). Particularly preferred electromagnetic radiation has a frequency of 10 kHz to 1 MHz.

The energy from the first energy source is used to specifically heat the particles without directly heating the liquid. This is achieved by using a material that has a higher absorption capacity than the surrounding liquid because of the energy transmitted by the first energy source. The energy absorbing particles are preferably selected from the group of solid materials that can absorb radiation in the above-mentioned range. Preferred materials are selected from the group of glass particles, for example metals or metal oxides, with an additional component for increasing the dielectric constant. More preferably, the metal is a glass with magnetic pigment distributed therein. Preferably the mass of the solid particles at at least 30%, more preferably at least 50%, is 2 pg to 20 ng, preferably 30 pg to 2 ng. In another embodiment for defining the particles, particularly in the case of substantially spherical particles, a particle size of at least 30%, more preferably 50%, is preferably 0.1-100 μm, more preferably 0.5- 10 μm. Preferably, the average diameter of the particles is 1-10 μm, more preferably 2-5 μm. In yet another aspect defining the particles of the present invention, at least 30%, more preferably 50% or more of the particles have a diameter relative to the volume of the particles of 2 × 10 ⁸ mm ⁻² to 2 × 10 ² mm ⁻² , More preferably, it has a ratio of 5 × 10 ⁵ mm ⁻² to 8 × 10 ⁴ mm ⁻² .

More preferably, the solid particles, 500~1000Jkg ^-1 K ^-1, more preferably the heat capacity of 600~800Jkg ^-1 K ^-1.

  More preferably, the solid particles are included in the mixture at 0.1 to 20 wt%, more preferably 0.5 to 5 wt%.

Preferably, the particles have the additional property of being able to bind to nucleic acids. This can be achieved either by non-specific adsorption or by specific binding. It is generally known that non-specific binding of nucleic acids occurs at the glass surface. Therefore, particles with a glass surface are preferred. Specific binding can occur when a nucleic acid having a sequence complementary to the nucleic acid is bound to the surface of the particle. Methods of attachment, particularly covalent attachment to the surface of oligonucleotides are well known.

  In one aspect of the invention, the solid particles have at least one primer having a first sequence that binds to these surfaces. Primers are compounds that can be extended at their 3 'ends with a polymerization agent and extension blocks, such as mononucleotides, using nucleic acids that bind to a template for sequencing.

  With reference to FIG. 1, the manner of binding of nucleic acids to particles is described. In the first embodiment shown in FIG. 1a, a first amount of first particles binds to the first primer, and a second amount of the same or different second particles binds it to the second primer. In the second embodiment shown in FIG. 1b, the particles bind both the first primer and the second primer. In the third embodiment shown in FIG. 1c, the first primer binds to the particles and the second primer is dissolved in the liquid.

  The present invention utilizes the induction energy of the first energy source in a pulse sufficient for heating the solid particles to 90 ° C. or more, 0.001 to 10 seconds, preferably 0.001 to 1 second. To do. Inducing energy during the pulse can be done by activating the energy source and inducing energy into the mixture for a period of time. The time is sufficient to heat the particles. The present invention uses the ability of solid particles to sequentially pass some heat to the liquid immediately surrounding the particles. The liquid heated by the particles depends on the length of the pulse. Generally longer pulses will increase the amount of liquid being heated, and shorter pulses will only heat a smaller amount of liquid. Furthermore, the heat transfer coefficient from the particles to the liquid will determine the heat provided to the liquid. A larger heat transfer coefficient delivers more heat. Furthermore, the flow characteristics of the liquid will determine the amount of heat delivered. The more convection is done in the liquid, the greater the amount of heat delivered, but the lower the temperature present in the liquid immediately surrounding the particles.

  Preferably, the layer heated to about 90 ° C. is about 100 nm thick in the present invention. The layers further away from the particles are heated but not reached that temperature. A temperature of 90 ° C. or higher is required in order to provide conditions for denaturation of double-stranded nucleic acid, ie, formation of a single strand. This is necessary to anneal the new primer to the extension product and the target nucleic acid.

  An essential characteristic of the present invention is that the temperature of the particles and the temperature of the liquid immediately surrounding the particles is 90 ° C. or higher. On the other hand, at an ambient pressure of 100 ° C., the temperature should not exceed the boiling point of the liquid. The energy required to heat the particles to that temperature can be determined by several experiments, changing the pulse length and measuring the temperature, and taking into account whether the liquid exhibits boiling bubbles. It can be measured easily.

  The method according to the invention has a different need for the embodiment when the primer is bound to the particle compared to the embodiment where the nucleic acid is immobilized directly on the particle surface. In the case of only immobilized primers (in the case of FIGS. 1a and 1b), a primer extension step on the surface is produced in the extension product bound to the particles, which is further hybridized with the original template. Are combined. Heating the particle surface and layer to a temperature of about 95 ° C. dehybridizes the template from the extension product. The mold floats from the surface to the liquid portion outside the layer. Only by reducing the temperature of the layer to a temperature below the melting point of the respective hybrid, the template can bind to nucleic acids, such as other immobilized primer molecules or complementary extension products, on the surface. If the particles are not heated (outside the energy pulse), this can occur over time.

  In embodiments where only one primer is bound to the surface, the extension product binds to the dissolved primer that has migrated to the surface after the extension product hybrid has denatured and the temperature has dropped to the annealing temperature. be able to. The primer is extended and the cycle can be repeated. The template can again bind to a surface-immobilized primer that can be extended under annealing conditions.

  In embodiments where no immobilized primer is used, the primer-template hybrid is formed in the liquid portion that is not in the layer and elongation occurs. As the extension product and template hybrids enter the layer, denaturation occurs and, after either the end of the heat pulse or floating outside the layer, the extension product and template hybridize with their respective primers. Can be soyed.

  The portion of the liquid that is not located in the layer around the particles is maintained at a temperature substantially lower than the temperature in the layer. Preferably, it is useful in embodiments of the invention where the primer is dissolved in the remaining portion of the liquid and the portion of the liquid outside the layer is maintained at the temperature required for primer annealing and / or extension. Thus, a preferred embodiment of the present invention comprises maintaining the liquid at a substantially constant temperature of 50-60 ° C.

  Thus, maintaining the temperature at a constant level requires heating and / or cooling the mixture, preferably the liquid. Whether a liquid requires heating or cooling depends on the amount of energy induced in the particles and further provided to the surrounding liquid by the particles. If only a small amount of energy is transferred to the liquid, it may be necessary to heat the liquid to an appropriate level. This can occur when there is low convection in the mixture (no mixing of particles in the mixture). Conversely, if the mixture is agitated, the mixture will need to be cooled to reach the desired temperature in the mixture.

  Means for heating the portion of the liquid not contained in the layer are known. These can preferably be selected from the group consisting of thermoelectric heating, resistance heating, and fluid mediated heating. Suitable heating sources are known, for example conventional thermal cyclers. A particularly preferred heat source is a resistance heater. In these, an electric current is used to heat a heater, which is a container containing the mixture, for example an aluminum block.

  Means for cooling are also known. These include, for example, liquid cooling by directing a gas stream or liquid to a container or a block containing the container, or by thermoelectric cooling realized, for example, with a Peltier element.

  The method according to the present invention more preferably includes both cooling and heating during the same amplification path if heating is required at the beginning when less heating is provided by heating the particles. In the second half of the process, the amount of heat transferred from the particles will not require independent heating of the liquid and will require further cooling. The problem can be easily eliminated by measuring the temperature in or near the liquid, eg the block containing the container, and by using sensors to control the temperature by a computer program.

  Thus, in a highly preferred embodiment, the present invention comprises two different heating modes and one cooling mode; the heating and cooling processes are controlled and regulated by a computer program that depends on the temperature of the liquid. Is done.

  In a broad sense, the present invention is a method for heating a mixture of liquid and particles comprising the steps of providing the mixture in a container and heating the mixture, wherein the mixture comprises: It relates to a method in which different types of heating sources are heated simultaneously and / or continuously.

  The container according to the invention is a container for holding the mixture under the conditions of the above method. Thus, the container is heat resistant, resistant to the amount and type of radiation provided to the mixture, resistant to the reagents contained in the mixture, and rigid so that the mixture does not escape from the container. Should.

  Preferably, the at least one heating method is selected from the group consisting of induction heating, thermoelectric heating, resistance heating, radiant heating, and fluid mediated heating.

  Induction heating is performed by application of electromagnetic radiation at a frequency / wavelength of 10 kHz to 1 MHz. Thermoelectric heating is accomplished by a heat source known as a Peltier element that generates heat by the well-known Peltier effect.

  Resistance heating utilizes the effect that the resistance of a small diameter wire in the current leads to energy loss due to heating.

  Radiant heating is created by directing radiation into the mixture in the container, the radiation having a wavelength that is absorbed by the mixture or container, and converting the radiation into thermal energy.

  Fluid mediated heating is accomplished by inducing a fluid, i.e., a liquid or gas stream, into a container or mixture, the fluid having a temperature above that of the mixture. The higher the difference in temperature between the fluid and the mixture ring, the faster the mixture is heated. Furthermore, the flow characteristics of the fluid will determine the amount of heat transferred.

  Preferably, the first heating is performed by non-contact heating, and the second heating is performed by contact heating. Contact heating is heating in which the heat medium is in contact with the material to be heated so that energy can flow through the contact surface between the heat medium and the material. If the particles in the mixture are dispersed in the mixture, heating to a temperature above the temperature of the liquid surrounding the particles of the mixture cannot use contact heating. However, heating of the liquid portion outside the layer around the particles can often be done by contact heating.

  If the energy source cannot be in direct contact with the material to be heated, non-contact heating is performed. Therefore, this mode of heating is preferred for heating the particles in the mixture without substantially directly heating the liquid portion outside the layer. The most preferred mode of non-contact heating is heating by radiation, for example heating by radiation that is not absorbed by the liquid part not in the surrounding layer of particles.

  Further heating modes can be distinguished by the heat transfer medium. Preferably, the liquid is heated by a combination of induction heating and resistance heating.

  Most preferably, the present invention relates to a method wherein the heating is performed using a combined heating source. Such a heat source can be applied to both contact heating as well as non-contact heating and is particularly suitable for applications where both methods are used simultaneously.

  The heating source according to the present invention is a resistance heater and an induction heating coil. A preferred design is a heating coil with a resistance defined for resistance heating that is designed to generate an electromagnetic field for induction heating as soon as operating with a suitable alternating current of a suitable frequency. The coil may be formed of wire or may be designed as a conductor of any kind of material in other ways, for example in a printed circuit board or on a substrate such as ceramic or polyimide. Another option is that the coil is formed on a suitable substrate by thin- or thick film technology. The coil may be located at the bottom, top, or side of the receptacle, or may enclose the device so that the device is in a coil that depends on the design of the coil. The coil should be designed to produce the strongest magnetic field possible. The coil may be part of an oscillating circuit (parallel resonant circuit) operated by a high frequency generator at the resonant frequency of the circuit to generate a strong magnetic field.

  Materials for receptacles heated by contact heating through combined heating elements need to have high thermal conductivity to achieve good heat transfer from the heater to the device including the chamber for the mixture . Furthermore, the material must be inert to the electromagnetic field, which means that it should not affect the electromagnetic field for non-contact heating in a negative manner. Preferably, the receptacle material is selected from the group of materials having zero electrical conductivity. More preferred materials are selected from the group of ceramic materials (eg AIN ceramic).

  For resistance heating, the heater needs to be in contact with a receptacle for the device containing the mixture in the chamber (contact heating). For induction heating of the particles and the surrounding layer of particles, the combined heater needs to be placed at a suitable distance relative to the device having the chamber containing the mixture (non-contact heating). The most preferred design is flat on the substrate located close enough to the chamber containing the mixture to be able to contact the receptacle for contact heating and to have the mixture in the center of the generated electromagnetic field for non-contact heating of the solid particles. Coil.

  Figures 2a and 2b show possible arrangements of heating elements combined in a device (3) with two coils (1) formed by wires, a receptacle (2) and a chamber.

  FIG. 3 shows, in an exploded view, a possible arrangement of combined heating elements, performed as a flat device (6) with a flat coil (4) and receptacle (5) on a suitable substrate and a chamber.

  In a preferred embodiment of the invention, similar to the situation described above for the method for nucleic acid amplification, the method further comprises active cooling of the liquid.

  Another aspect of the invention is an apparatus for heating one or more liquids comprising a receptacle for receiving a container comprising a chamber for containing a liquid, a first heat source, and a second heat source, The first and second heat sources, when placed in the receptacle, are located in the instrument to be effective for heating the contents of the chamber of the container.

  Another aspect of the present invention is a system for heating a liquid comprising a container comprising one or more chambers for containing a liquid and one heat source and a second heat source, wherein said heat source Is positioned in the system to be effective for heating the contents of the chamber of the device when placed in a receptacle for the container.

  Such a system is useful for the methods disclosed above. Conveniently, the system has a housing comprising the heat source, particularly preferably the heat generated by the heat source is mainly guided to the container and separated so that it is not induced to the surroundings. This is particularly important for radiation sources or for health reasons.

  As described above, a system in which the first heat source is a non-contact heat source and the second heat source is a contact heat source is preferred.

Example 1
Treatment of liquids containing solid particles by radiation A model experiment with a mixture of the following components was created and performed in the laboratory:

  In the model experiment, two samples were heated from room temperature to a final temperature of about 100 ° C. by induction heating in a polypyrene tube. The tube was placed in a 3-turn induction heating coil. The experiment was performed using different power sources as energy sources operating at different frequencies. The best results were obtained with a power supply operating at about 200 kHz and providing 20 kW of power. The power was continuously operated during the experiment until the sample reached the final temperature. The temperature was measured with a thermocouple.

The following table gives a summary of the results from the experiment.

  The experiment shows that it is generally possible to heat solid particles in an appropriate volume of aqueous solution suitable for analysis of samples in medicine by non-contact induction heating. In this experiment, the entire liquid in which the solid particles were suspended was heated to the final temperature, so more energy was required in the experiment than if only the solid particles and the surrounding layer had to be heated to the final temperature. It was said.

Example 2
Theoretical Experiment / Model Simulation In the sample model, the energy Q _in entering the single particle via non-contact heating and the resulting energy loss Q _out to the periphery of the cooler liquid layer were simulated (see FIG. 5). ).

  The model was set up on a single bead. The first input to the model is the total power provided by non-contact heating. The total power is divided later through the calculated number of particles to obtain input per single bead. The second input is the particle size that affects the number of particles, as well as the surface area of the solid particles, for a given total mass of solid particles. The third input value is the total mass of the particles. The number of particles is calculated as follows:

により計算される。 The mass of a single particle is

Is calculated by

によりもたらされる。 The volume of one particle is

Brought about by.

  The model calculates a given time interval Δt, for example, a step of 0.001 seconds per step.

により計算される。 The beads are regarded as a container with an energy input Q _in and an energy output Q _out . The change in energy from time t-1 to time t is

Is calculated by

The input energy from induction heating is a product derived from the input power and time interval.

により計算される。 The energy of the beads at time t is

Is calculated by

によりもたらされ、以下のとおり実際のビーズ温度に加えられる。

The change in bead temperature from time t-1 to time t is

And is added to the actual bead temperature as follows:

である。 On the output side, energy loss due to heat transfer to the liquid layer is

It is.

The temperature T _liquid of the surrounding liquid layer was considered to remain stationary at a temperature of 50 ° C. In order to satisfy the hypothesis, the surrounding liquid layer must be cooled by appropriate means. At the start of the simulation, the beads have the same temperature as the liquid layer.

  In addition to the results from model experiments conducted in the laboratory, all inputs were fed into the model. While varying the size of the solid particles and the total amount of solid particles in the solution, the temperature that can be theoretically achieved with a given input can be calculated.

Results The following paragraph summarizes the results obtained from the simulation based on the model described above. FIG. 4 shows the dependence between the particle size and the power required to obtain a fixed size 5 mg solid particles heated to a temperature of 95 ° C. in water kept at a constant temperature of 50 ° C. .

  The only required power shown in FIG. 4 is the power that the beads must be bound to reach the desired temperature. The total power required by the system to achieve the same effect depends on the energy efficiency of the entire system. The generator for the electromagnetic field in Example 1 provided 20 kW of power, resulting in a binding power of about 70 W to the system heating the beads and the liquid in which they were suspended.

Three possible methods of immobilized primer on solid particles are shown (1a, 1b, and 1c). Two embodiments are shown for placing the heating coil in the vicinity of the chamber in the apparatus (FIGS. 2a and 2b). A very flat device with a flat chamber is shown. Figure 4 shows the dependence of the particle size versus the power required to reach a particle temperature of 95 ° C with 5g total mass particles in water kept constant at 50 ° C. 2 illustrates the basic principle of the present invention, exemplified in a single particle.

Explanation of symbols

DESCRIPTION OF SYMBOLS 1 Heating wire 2 Receptacle 3 Apparatus with a chamber 4 Flat coil 5 Receptacle 6 Flat apparatus with a chamber

Claims (20)

A method for nucleic acid amplification comprising:
(A) providing a mixture of liquid containing solid particles having a higher first energy source absorption capacity than the nucleic acid and the liquid, the mixture containing a reagent necessary for amplifying the nucleic acid;
(B) a step of introducing the first energy in a pulse of 0.001 to 10 seconds for a time sufficient to heat the solid particles to a temperature of 90 ° C. or higher;
Comprising
A method wherein the liquid is actively maintained at annealing and elongation temperatures.   The method of claim 1, wherein the particles have an average diameter of 0.1 to 100 μm.   The method according to claim 1, wherein the solid particles are bonded solid particles.   The method according to claim 1, wherein the first energy source is an electromagnetic field.   The method according to any one of claims 1 to 4, wherein the energy is radiant energy having a frequency of 1 kHz to 50 GHz.   The method according to claim 1, wherein a mass of at least 30% of the solid particles is 2 pg to 20 ng. The method according to claim ¹ , wherein the solid particles have a heat capacity of 500 to 1000 Jkg ⁻¹ K ⁻¹ .   The method according to any one of claims 1 to 7, wherein the solid particles are contained in the mixture at 0.1 to 20 w-%.   9. A method according to any one of the preceding claims, wherein the solid particles have primers that bind to their surfaces.   10. A method according to any one of the preceding claims, further comprising maintaining the liquid at a substantially constant temperature of 50-60C.   Providing a mixture of said liquid comprising nucleic acids to be amplified and solid particles having a higher first energy source absorption capacity than the liquid in the container, and heating the mixture, for heating the liquid Method, characterized in that the liquid is heated simultaneously and / or continuously by different types of heat sources. At least one heating method comprising:
-Induction heating,
-Thermoelectric heating,
-Resistance heating,
-Radiant heating, and
-Fluid mediated heating,
12. A method according to any one of claims 1 to 11 selected from the group consisting of:   The method according to claim 1, wherein the mixture is heated by a combination of induction heating and resistance heating.   The method of claim 13, wherein the first heating is performed by contact heating and the second heating is performed by non-contact heating.   15. A method according to any one of claims 1 to 14, wherein the heating is performed using a combined heat source.   16. A method according to any one of the preceding claims, wherein the method comprises inductively heating the solid particles dispersed in the liquid.   17. A method according to any one of the preceding claims, further comprising active cooling of the mixture. A system for heating a mixture comprising one or more nucleic acids to be amplified and a liquid comprising solid particles having a higher first energy source absorption capacity than the liquid comprising:
An apparatus comprising one or more chambers for containing the mixture, and
-A first type of heat source; and
-A second heating source;
Comprising
Wherein the heating source is located in the system to be effective to heat the contents of the chamber of the device when placed in a receptacle for the device.   The system of claim 18, wherein the first heating source is a contact heating source and the second heating source is a non-contact heating source. An apparatus for heating one or more mixtures each comprising a nucleic acid to be amplified and a liquid each comprising solid particles having a higher first energy source absorption capacity than the liquid, comprising:
A receptacle for receiving a device comprising a chamber for containing the mixture;
-A first heating source, and
A second heating source,
Comprising
Wherein the device is positioned in the device to be effective for heating the contents of the chamber of the device when the first and second heat sources are placed in the receptacle.

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